Neuron-specific enolase and malignant lymphomas (23 cases)

Summary The immunoreactivity of polyclonal antiserum to neuron-specific enolase (NSE) has been investigated. Twenty-three cases of malignant lymphoma (ML) were studied and compared with previously published reports. In our study 11 out of 23 cases showed strong or weak NSE positivity; any type of ML could be positive or negative even among B or T cell ML. This study indicated that polyclonal NSE is not a specific marker; it might be an inconstant marker of ML with no apparent correlation between reactivity and morphology or phenotype.     

Novel pluripotential neural progenitor lines exhibiting rapid controlled differentiation to neurotransmitter receptor‐expressing neurons and glia

The immortalization of progenitor cells from embryonic murine hippocampus using oncogene‐carrying retroviral vectors is described. Use of a vector encoding the oncogene v‐myc results in lines of nestin‐positive progenitor cells. Limited differentiation ensues if the cells are cultured in the presence of dibutyryl cyclic adenosine monophosphate. In contrast, use of a vector in which the extracellular portion of the epidermal growth factor (EGF) receptor is fused to the neu tyrosine kinase generates lines of pluripotential nestin‐positive progenitor cells, which differentiate upon withdrawal of EGF into neurons and glia. Differentiated neurons expressing action potentials and neurotransmitter receptors make up a high proportion of the cells. These cell lines are useful tools to investigate the characteristics of differentiating neurons and glia, as well as to screen neuroactive drugs. This work has been reported in preliminary form as an abstract (1996 Society for Neuroscience Abstract, #606.20, p. 1537).     

Studies on mutants deficient in the photosystem I reaction centers in Chlamydomonas reinhardtii

Summary Genetic analysis of 25 nuclear mutants defective in the chlorophyll-protein complex CP1 was undertaken. The mutants belong to 13 complementation groups scattered throughout the nuclear genome. All these mutants lack the apoprotein of CP1 and, in addition, a specific set of six low molecular weight thylakoid polypeptides. System I particles obtained by treating WT thylakoid membranes with detergent specifically contain those polypeptides which the mutants lack. These observations suggest that a particular sub-structure of the thylakoid membrane associated with the photosystem I activity is missing from all 25 mutants studied, and that this general phenotype can result from mutation at any one of several unlinked Mendelian loci.     

Analysis of very long-chain fatty acids using electrospray ionization mass spectrometry

Elevated levels of very long-chain fatty acids (VLCFA) in plasma and tissues are the biochemical hallmark for patients with X-linked adrenoleukodystrophy (X-ALD). Current methods for the determination of VLCFA levels are laborious and time-consuming. We describe a rapid and easy method using electrospray ionization mass spectrometry (ESI-MS) with deuterated internal standards. VLCFA are hydrolyzed, extracted, and quantified in less than 4h. This includes 2h of hydrolysis and 4min of quantification. We validated the method by analyzing 60 plasma samples from controls and patients with X-ALD or Zellweger syndrome using both the ESI-MS protocol and an established method for VLCFA analysis using gas chromatography (GC). The C26:0 concentrations determined with ESI-MS in plasma and fibroblasts of X-ALD patients are in good agreement with those reported previously for GC and GC-MS. Besides saturated straight chain VLCFA, we also determined the concentrations of the mono-unsaturated VLCFA C24:1 and C26:1 and established that while C24:1 levels are not elevated, C26:1 levels are elevated in both plasma and fibroblasts from X-ALD patients.     

P67L: a cystic fibrosis allele with mild effects found at high frequency in the Scottish population.

Only three mutant cystic fibrosis (CF) alleles have to date been established as conferring a dominant mild effect on affected subjects who are compound heterozygotes. We now add a fourth, P67L, which occurs on about 1.4% of Scottish CF chromosomes. Among 13 patients (12 unrelated) with this allele, the average age at diagnosis was 22.5 +/- 11.3 years. None of the cases had consistently raised sweat chloride concentrations, the average value being 57 +/- 9 mmol/l; 77% of the patients were pancreatic sufficient. When compared to three other established mild CF alleles, R117H, A455E, and 3849 + 10kb C-T, a compound heterozygote for P67L has minimal disease and clinical suspicions are unlikely to be confirmed other than by DNA typing.     

Glutamine synthetase is essential in early mouse embryogenesis.

Glutamine synthetase (GS) is expressed in a tissue-specific and developmentally controlled manner, and functions to remove ammonia or glutamate. Furthermore, it is the only enzyme that can synthesize glutamine de novo. Since congenital deficiency of GS has not been reported, we investigated its role in early development. Because GS is expressed in embryonic stem (ES) cells, we generated a null mutant by replacing one GS allele in-frame with a beta-galactosidase-neomycine fusion gene. GS(+/LacZ) mice have no phenotype, but GS(LacZ/LacZ) mice die at ED3.5, demonstrating GS is essential in early embryogenesis. Although cells from ED2.5 GS(LacZ/LacZ) embryos and GS(GFP/LacZ) ES cells survive in vitro in glutamine-containing medium, these GS-deficient cells show a reduced fitness in chimera analysis and fail to survive in tetraploid-complementation assays. The survival of heavily (>90%) chimeric mice up to at least ED16.5 indicates that GS deficiency does not entail cell-autonomous effects and that, after implantation, GS activity is not essential until at least the fetal period. We hypothesize that GS-deficient embryos die when they move from the uterine tube to the harsher uterine environment, where the embryo has to catabolize amino acids to generate energy and, hence, has to detoxify ammonia, which requires GS activity.